For analyzing biological and/or chemical activities of substances or particles and/or molecules present in the substances, data of the samples to be analyzed are acquired. For acquiring data, microscopes, particularly confocal microscopes, are used. In doing so, certain components of a sample are marked with fluorescent markers, where it is possible to draw conclusions with respect to reactions within the sample on the basis of the emission of fluorescence of these markers.
The reaction of a cell to a particular substance can be detected, for example, by the migration of a molecule marked with a fluorescent marker, for example, into the nucleus. An analyzing method suitable therefor is described in U.S. Pat. No. 5,989,835. In this method, the nucleus of a cell is marked or colored in a first step. The coloring is performed with a fluorescent marker that may be stimulated by a UV laser. Further, a transcription factor in the cytoplasm surrounding the nucleus is marked with another fluorescent marker. By stimulating the nucleus with a UV laser and a threshold value method, a mask separating the nucleus from the cytoplasm is prepared. Subsequently, the mask is reduced in size so that it is guaranteed that a substantially circular first portion is exclusively arranged within the nucleus. In the next step, the mask border is enlarged so that a ring is created which exclusively lies in the cytoplasm. By comparing the two cell portions, for example, it can be detected that an active substance marked with a color marker has migrated into the nucleus, since the luminosity of the two portions changes in this case.
First, this method described in U.S. Pat. No. 5,989,835 has the disadvantage that the nucleus has to be located. This is only possible by using special colors or special color markers since it has to be ensured that they react with the nucleus to be able to detect the position of the nucleus. The color markers used in this case, Hoechst 33342 and 33258, for example, can only be stimulated by a UV laser. The acquisition of UV lasers, however, is expensive, and they have high operational costs due to their high cooling water consumption. Another disadvantage of the afore-described method consists in that meaningful results can only be achieved if the ring is arranged completely within the cytoplasm. This is not the case, for example, when the nucleus is arranged in the border portion of the cytoplasm. Another disadvantage of the afore-described method consists in that always a definitely defined surface in the form of the nucleus is defined and detected. Thus, the method is expensive and inflexible.
It is the object of the invention to provide an analyzing method for chemical and/or biological samples where the trouble with detecting changes of the sample is reduced.
This object is solved, according to the invention, by the method for analyzing chemical and/or biological samples, particularly with high-throughput or medium-throughput screening installations, with the steps of: respectively producing at least one particle image (42) of at least one sample with at least one particle being included in the sample, respectively; (b) defining several particle zones (14,18; 22,24; 32,34,36) independent of subparticular compartments; (c) acquiring particle data of the sample independently of the zone; and (d) evaluating the acquired particle data.